Several reports have shown that electrical and ionic changes occurring in acute myocardial ischemia can be closely mimicked by exposure of tissue to hypoxic, acid-, and glucose-free solutions at elevated [K+]o. In the present work, this approach was chosen to distinguish between the combined effects of hypoxia, substrate withdrawal, and acidosis, and the effects of two different levels of [K+]o (4.7 mM and 11.5 mM) on intracellular sodium activity and resting membrane potential. Measurements were made with microelectrodes in isolated guinea pig papillary muscles. In normoxia at 4.7 mM [K+]o, intracellular sodium activity was 7.5 mM (+/- 1.9 mM, SD) during stimulation at 1 Hz. Combined hypoxia, substrate withdrawal, and acidosis increased intracellular sodium activity significantly, by 3-4 mM in 4.7 mM [K+]o and by approximately 2 mM in 11.5 mM [K+]o, after 9-10 minutes. Increasing [K+]o in normoxic solution decreased intracellular sodium activity by 1.9 mM (+/- 1.3 mM, SD). The transition from normal (4.7 mM [K+]o) Tyrode's solution to "ischemic solution" (hypoxia, acidosis, substrate withdrawal, 11.5 mM [K+]o) was associated with a small initial increase and a subsequent decrease of intracellular sodium activity. The steady state level after 12 minutes was not significantly different from the level in normal Tyrode's solution. The secondary decrease of intracellular sodium activity coincided with the gradual development of inexcitability and was absent in quiescent preparations. Combined hypoxia, acidosis, and glucose-withdrawal produced a depolarization by 7-10 mV at 4.7 mM and at 11.5 mM [K+]o, probably reflecting cellular potassium loss and extracellular potassium accumulation in the restricted extracellular space.(ABSTRACT TRUNCATED AT 250 WORDS)